Mixed lineage kinases (MLKs) are a family of serine/threonine protein kinases that function in a cascade of phosphorylation reactions to control the activity of specific mitogen activated protein kinases (MAPKs). Gallo and Johnson, Nature Reviews; 3: 663-664 (2002). MLKs act as MAPK-kinase kinases (MAPKKKs) to activate, for example, the stress activated protein kinase/c-Jun N-terminal kinase (SAPK/JNK) and p38 kinase pathways, through phosphorylation of MAPK kinases (MKKs). Bloem et al., J. Mol. Cell. Cardiol.; 33: 1739-1750, (2001). SAPK/JNK and p38 kinase are specific MAPKs that, like other eukaryotic MAPKs, are activated in response to a multitude of stimuli including exposure to inflammatory cytokines, hormones, and growth factors as well as cellular stresses such as heat shock, inhibition of protein glycosylation, and exposure to ultraviolet irradiation. Fanger et al., Curr. Opin. Genet. Dev.; 7(1): 67-74 (1997).
Three subfamilies of MLKs have been previously identified and grouped on the basis of domain arrangements and sequence homology within their catalytic domains. The MLKs (MLK1-MLK4) contain an amino-terminal SRC homology domain (SH3), followed sequentially by a catalytic kinase domain, a leucine zipper region, and a Cdc42/Rac-interactive binding domain (CRIB Motif). MLK1-MLK4 share approximately 75% homology within their catalytic domains and approximately 65% homology from the SH3 domain to the CRIB motif. However, the carboxy terminus of each of these kinases diverge, indicating these regions may serve different regulatory functions. Nature Reviews; 3: 663-664 (2002). The duel leucine zipper bearing kinases (DLKs) represent the second family of MLKs and are characterized by a kinase domain followed by two leucine zipper motifs, separated by a 31 amino acid spacer. The catalytic domains of the two DLKs (DLK and leucine zipper kinase (LZK)) share approximately 87% sequence homology but again diverge in their carboxy terminus. The final subfamily of MLK is represented by zipper sterile α αmotif kinase (ZAK). ZAK shares homology with the other MLKs through the leucine zipper domain, but again diverges from the others at the carboxy terminus. Nature Reviews; 3: 663-664 (2002). While all of the MLKs have been shown to activate the c-Jun N-terminal kinase pathway, some have also been shown to activate the p38 kinase pathway as well. In addition, other MAPKKKs have been identified which activate the JNK and p38 kinase pathways, including MEK kinase; apoptosis inducing kinase 1 (ASK1); and transforming growth factor beta (TGFβ)-activated kinase (TAK1). Nature Reviews; 3: 663-664 (2002).
Congestive heart failure (CHF) is a complex disorder with several etiologies including hypertension, myocardial injury, and hemodynamic overload. One of the adaptive responses of the heart to these stresses is hypertrophy of the cardiac myocyte, characterized by altered gene transcriptional regulation, increased protein synthesis, and increased organization of the myofibril. This hypertrophy, in turn, may lead to remodeling of the heart and subsequent failure. J. Mol. Cell. Cardiol; 33: 1739-1750, (2001). Evidence suggests that activation of signal transduction pathways, including MAPK and stress activated protein kinase (SAPK/JNK) pathways contribute to hypertrophic changes in the heart. (See generally: Sugden et al., J. Mol. Med.; 76: 725-746, (1998); Force et al., Gene Expression; 7: 337-348, (1999); Ramirez et al., J. Biol. Chem.; 272: 14057-14061, (1997); Bogoyevitch; Cardiovasc. Res., 45: 826- 842, (2000); Hines et al.; J. Mol. Cell. Cardiol., 30: 485-494, (1998) and Clerk et al.; Am. J. Cardiol., 83: 64H-69H, (1999)).
Recently, a novel MAPKKK, designated as MLK-7, was identified from a database mining effort of a cDNA library from human failed heart tissue. The cDNA encodes for a 55 kDa. protein with serine/threonine kinase activity when expressed and purified from insect cells. J. Mol. Cell. Cardiol; 33: 1739-1750, (2001) In addition, MLK-7 activates the SAPK/JNK1 pathway in rat neonatal cardiac myocytes and modulates fetal expression of marker genes for cardiac hypertrophy. Specifically, MLK-7 increased expression of atrial natruiretic factor (ANF) and decreased expression of α myosin heavy chain (αMHC) mRNAs in rat neonatal cardiac myocytes. Furthermore, MLK-7 also increased protein synthesis in cardiac myocytes as evidenced by increased [14C] phenylalanine incorporation in MLK-7-infected cells. J. Mol. Cell. Cardiol; 33: 1739-1750, (2001). Taken together this data suggests that MLK-7 is implicated in the hypertrophy of cardiac myocytes that occurs in response to the various etiologies of congestive heart failure. Thus antagonists of MLK-7 activity may have utility in the treatment patients suffering from CHF.
Surprisingly, and in accordance with this invention, Applicants have discovered a series of dihydropyrrolopyrazole compounds of Formula I which are believed to be novel, as more fully described below, which are potent antagonists of the recently identified MLK-7. Such antagonists could be useful for the treatment of CHF and could therefore address a long felt need for a safe and effective treatment for CHF. The treatment of cardiovascular disorders is hereby furthered.